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Biomedical subjects

L Garcia

Publications and source records attributed to L Garcia.

At least 19 recordsLinked to original sources
PubMed2012

MRI roadmap-guided transendocardial delivery of exon-skipping recombinant adeno-associated virus restores dystrophin expression in a canine model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) cardiomyopathy patients currently have no therapeutic options. We evaluated catheter-based transendocardial delivery of a recombinant adeno-associated virus (rAAV) expressing a small nuclear U7 RNA (U7smOPT) complementary to specific cis-acting splicing signals. Eliminating specific exons restores the open reading frame resulting in translation of truncated dystrophin protein. To test this approach in a clinically relevant DMD model, golden retriever muscular dystrophy (GRMD) dogs received serotype 6 rAAV-U7smOPT via the intracoronary or transendocardial route. Transendocardial injections were administered with an injection-tipped catheter and fluoroscopic guidance using X-ray fused with magnetic resonance imaging (XFM) roadmaps. Three months after treatment, tissues were analyzed for DNA, RNA, dystrophin protein, and histology. Whereas intracoronary delivery did not result in effective transduction, transendocardial injections, XFM guidance, enabled 30&#xb1;10 non-overlapping injections per animal. Vector DNA was detectable in all samples tested and ranged from <1 to >3000 vector genome copies per cell. RNA analysis, western blot analysis, and immunohistology demonstrated extensive expression of skipped RNA and dystrophin protein in the treated myocardium. Left ventricular function remained unchanged over a 3-month follow-up. These results demonstrated that effective transendocardial delivery of rAAV-U7smOPT was achieved using XFM. This approach restores an open reading frame for dystrophin in affected dogs and has potential clinical utility.

Animals↗
PubMed1995

Myotube driven myogenic recruitment of cells during in vitro myogenesis.

Muscular dysgenesis (mdg) is a recessive lethal mutation in the mouse which drastically affects skeletal muscle development during embryonic life. Physiologically, the disease is characterized by a complete paralysis resulting from a lack of excitation-contraction coupling. Existing electrophysiological, biochemical, and genetic evidence shows that mdg/mdg mice express a basic alteration of L-type voltage-sensitive Ca2+ channels in skeletal muscle. Studies on mdg/mdg myotubes in primary culture have shown that +/+ fibroblasts or +/+ Schwann cells may fuse with them and correct their functional deficiency by genetic complementation. As the spontaneous formation of heterocaryons is thought to be an exclusive property of myoblasts, we asked whether fibroblasts may have changed their properties before fusion occurred. We used primary cells issued from sciatic nerves dissected from newborn transgenic mice carrying the pHuDes1-nls-LacZ transgene (Des-LacZ cells) as non-muscle cells. These cells were mainly fibroblasts (80%) positive for Thy1.1 and Schwann cells positive for S100. The cultures were negative for myogenic markers (desmin, troponin T), did not form myotubes long-term, and did not display significant activation of the muscle reporter gene (pHuDes1-nls-LacZ). After a few days in coculture with dysgenic or normal myotubes, the muscle reporter gene (beta-galactosidase) was detected both within dysgenic myotubes, correlating with the restoration of normal contractile activity, and normal myotubes. As well as confirming that fusion takes place, this shows that Des-LacZ cells nuclei incorporated into recipient myotubes express their own myogenic genes. Moreover, individual mononucleated Des-LacZ cells expressing beta-galactosidase were observed, indicating that myogenic genes were being expressed before fusion. This suggests a mechanism of myotube driven myogenic recruitment of cells during the in vitro myogenesis. Analysis of the distribution of the induced Des-LacZ cells (positive for beta-galactosidase) in compartmentalized muscle cocultures showed that in the presence of dysgenic myotubes, these cells were equally distributed in both myotube free and enriched areas, whereas in the presence of normal myotubes, the positive cells remained in close vicinity of the myotubes. This difference could be explained by the fact that the dysgenic phenotype might include release of the induction process from its normal controls. Our results are consistent with the idea of a transcellular mechanism triggering myogenic differentiation in non-muscle cells, and that myotubes themselves are able to drive myogenic recruitment of cells during the in vitro myogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Recording of a large-conductance chloride channel in normal rat lactotrophs.

Membrane current fluctuations resembling channel openings and closings were observed in the whole cell configuration of the patch-clamp technique in normal rat lactotrophs in primary culture. Using high-gain head stage in whole cell configuration, we characterized the nature and pharmacological properties of the ionic channel underlying these fluctuations. This channel, found in small numbers (< 10 per cell), was specific for Cl- because its reversal potential varied with Cl- gradients, according to the Nernst equation, and its unitary amplitude was linearly related to membrane potential from -100 to 0 mV. Slope conductance was close to 100 pS. Analyzing open times, we demonstrated its Ca2+ and potential dependence. Four sublevels were observed. We suggest that this channel, belonging to the background Cl- channel group, takes part in the regulation of intracellular Cl- concentration of normal rat lactotrophs.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1993

The relative anticaries effectiveness of sodium monofluorophosphate and sodium fluoride as contained in currently available dentifrice formulations.

A 36-month double-blind clinical caries study was conducted to determine the comparative anticaries efficacy of sodium monofluorophosphate (MFP) and sodium fluoride (NaF) as incorporated in dentifrice formulations. The test dentifrices which were utilized are commercially available in the United States and all have been granted the American Dental Association Seal of Acceptance for anticaries efficacy. The study was conducted in accordance with the 1988 Guidelines of the American Dental Association Council on Dental Therapeutics. The two formulations used in the NaF versus MFP comparison were Colgate Winterfresh Gel with 0.76% sodium monofluorophosphate in a silica base and Crest Regular Flavor Toothpaste with 0.24% sodium fluoride in a silica base. A third commercially-available NaF dentifrice, Colgate Junior Toothpaste with 0.243% sodium fluoride in a silica base, was included in the study for further comparison. Schoolchildren residing in Maine, U.S.A. received a baseline caries clinical examination and then were randomly assigned to the twice-daily use of one of the three dentifrices which were purchased in the marketplace and repackaged in plain white tubes. Caries clinical examinations were subsequently conducted after 2 and 3 years' use of the dentifrices. A total of 2,222 children participated in both the 2 and 3-year caries clinical examinations. The 36-month DFS (decayed and filled surfaces) and the DFT (decayed and filled teeth) mean caries increments for the three dentifrice groups were virtually identical. The children using the MFP formulation experienced a DFS of 1.50 and a DFT of 1.08; those using the Crest NaF formulation experienced a DFS of 1.48 and a DFT of 1.07; subjects in the third group (Colgate Junior Toothpaste, 0.243% NaF) had a DFS of 1.49 and DFT of 1.06. The statistical evaluation of the data consisted of the calculation of 90% confidence bounds for the ratio of (true) mean increments, in accordance with the 1988 American Dental Association Guidelines. Expressed in the language of the Guidelines, the analyses indicated that, (1) the anticaries efficacy provided by the MFP formulation, (Colgate Winterfresh Gel with 0.76% MFP) is "as good as" that provided by the NaF formulation, (Crest Regular Flavor Toothpaste, with 0.243% NaF), and (2) the anticaries efficacy provided by the one NaF formulation (Colgate Junior Toothpaste, with 0.243% NaF) is "as good as" that provided by the other, (Crest Regular Flavor Toothpaste, with 0.243% NaF).

American Dental Association↗
PubMed1992

Systemic pilocarpine increases deposition of and decreases responsiveness to inhaled carbachol in sheep.

The purpose of this study was to determine whether excessive airway secretions could serve as a barrier function against inhaled particulate matter. To increase airway secretions, six conscious sheep were treated with pilocarpine (0.8 mg/kg i.v.). Pilocarpine increased pulmonary resistance (RL) and total aerosol deposition within five breaths (AD5) as determined by the rebreathing of an inert monodisperse aerosol. When RL had returned to baseline, AD5 remained elevated [21 +/- 2% (SE), P < 0.05] and tracheal secretions were increased (237 +/- 77%, P < 0.05) above the values before pilocarpine administration. A carbachol aerosol dose-response curve was carried out at this time and compared with a control carbachol dose-response curve by calculating the dose of carbachol required to increase RL by 400% (PD400). Mean PD400 was increased postpilocarpine by 53 +/- 18 (P < 0.05) and 85 +/- 25% (P < 0.05) when normalized for increased aerosol deposition. Thus, pilocarpine decreased airway responsiveness to inhaled carbachol despite increasing aerosol deposition. The pilocarpine-induced airway hyporesponsiveness to inhaled carbachol is consistent with the hypothesis that excessive secretions have a protective role in the airways.

Administration, Inhalation↗
PubMed1992

Increased intracranial pressure in Alexander disease: a rare presentation of white-matter disease.

A patient with biopsy-proven Alexander disease presented with progressive head enlargement and clinical evidence of increased intracranial pressure. Lumbar puncture under sedation confirmed increased intracranial pressure with otherwise normal cerebrospinal fluid values. The association of megalencephaly, increased intracranial pressure and neuroimaging evidence of leukoencephalopathy should alert pediatric neurologists to the possibility of Alexander disease.

Biopsy↗
PubMed1992

N-cadherin and N-CAM in myoblast fusion: compared localisation and effect of blockade by peptides and antibodies.

The expression and distribution of two cell adhesion molecules, N-cadherin and N-CAM, at the surface of cultured leg muscle cells from 11-day-old chicken embryos were studied and compared. N-cadherin, which was expressed by fusing myoblasts, was down-regulated on old myotubes while N-CAM was still present. Both molecules, as viewed by confocal microscopy, appeared to have coaccumulated at the areas of contact between fusing myoblasts. However, immunogold electron microscopy did not reveal significant colocalization of N-cadherin and N-CAM, and their segregation after antibody-induced patching suggested the absence of direct interactions between N-cadherin and N-CAM. The role of the Ca2+ dependent cell adhesion molecule N-cadherin in myogenesis was investigated. Myoblast fusion was inhibited (1) with a synthetic peptide containing the H-A-V sequence and (2) with a monoclonal anti-N-cadherin antibody, demonstrating that N-cadherin-mediated cell adhesion is required for myoblast fusion. Under the same conditions no effect of anti-N-CAM antibodies was observed. Taken together these observations suggest that N-cadherin, acting independently from N-CAM, is a major cell adhesion molecule involved in embryonic myoblast fusion in vitro.

Amino Acid Sequence↗
PubMed1992

Leishmania chagasi antigens recognized in cured visceral leishmaniasis and asymptomatic infection.

Active visceral leishmaniasis is associated with antigen-specific immuno-suppression. However, cured patients develop a cellular immune response associated with resistance to reinfection. Recent studies have identified patients with asymptomatic or subclinical infections, which are also accompanied by an immune response. In order to identify subjects immune to Leishmania chagasi, we performed a skin-test survey in an endemic area in eastern Venezuela. The delayed-type hypersensitivity (DTH) response was assessed in patients cured of visceral leishmaniasis, as well as in their relatives and neighbors. Of the latter, 36 (34.2%) of 105 were positive and 26 (24.7%) of 105 gave intermediate responses. The DTH reaction correlated with age. The antigens recognized by a subgroup of cured patients, those with positive skin-test results, and controls (skin-test negative) were assessed by Western blotting with sera, and T cell immunoblotting with peripheral blood mononuclear cells. No consistent differences between the groups were noted in Western blots with L. chagasi antigens. T cell blots were performed on five patients from each group. For the cured patients and skin-test positive contacts, a significant proliferative response to fraction 12 (less than 20.5 kDa) was noted in four of five patients in each group. Cells from three of five cured patients and two of five skin-test-positive patients proliferated in response to fraction 4 (73-115 kDa). The response to other fractions was variable, with only a minority of patients responding to any one fraction. These data suggest that the antigens recognized by patients with evidence of immunity to L. chagasi are quite variable.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1991

Developmental modulation of physicochemical variants of the tailed asymmetric (16S) acetylcholinesterase by neuromuscular activity and innervation in the mouse embryo.

We have studied the physicochemical properties of acetylcholinesterase (AChE) during embryonic development of normal and functionally impaired mouse skeletal muscle, focusing on the tailed asymmetric (16S) form of the enzyme. The muscle-specific 16S AChE exists in two different variants. One is associated with extracellular matrix and is high-salt soluble (HSS, also termed hydrophilic AChE), whereas the other form is anchored to cell membranes and is detergent extractable (DE, or hydrophobic AChE). Before innervation during normal embryonic development, both hydrophilic and hydrophobic 16S AChE exist in equal amounts. After muscle innervation, there was an increase (amounting three-fold on E18) in the levels of hydrophilic vs. hydrophobic 16S AChE. This alteration of the relative proportions of the two variants of 16S AChE did not occur in chronically inactive muscles either from the mouse mutant, muscular dysgenesis, or from tetrodotoxin-treated mouse embryos. Taken together with previous reports, the present results suggest that postsynaptic membrane depolarization-induced Ca2+ fluxes are important in modulating not only the synthesis of 16S AChE, but also the relative proportions of both physicochemical variants of this molecular form of AChE.

Acetylcholinesterase↗
PubMed1991

Phosphatidylinositol is involved in the attachment of tailed asymmetric acetylcholinesterase to neuronal membranes.

1. We analyzed the mode of attachment of 16 S tailed acetylcholinesterase (AChE; EC 3.1.1.7) to rat superior cervical ganglion (SCG) neuronal membranes. Using extractions by high-salt (HS) and nonionic detergent (Triton X-100), we found two pools of 16 S AChE. 2. The detergent-extracted (DE) 16 S AChE was tightly bound to membranes through detergent-sensitive, high-salt insensitive interactions and was distinct from high-salt-soluble 16 S AChE. The detergent-extracted (DE) 16 S AChE constituted a significant proportion of about one-third of the total 16 S AChE. 3. Treatment of the neuronal membranes by a phosphatidylinositol-specific phospholipase C (PIPLC) resulted in the release of some, but not all DE 16 S AChE, indicating that a significant amount of the neuronal DE 16 S AChE, about one-third, is anchored to membranes through a phosphatidylinositol containing residue. Thus, a covalent association of a glycolipid and catalytic or structural AChE polypeptidic chains occurs not only for dimeric AChE but also for the asymmetric species of AChE. 4. The complex polymorphism of AChE is due not only to different globular or asymmetric associations of catalytic and structural subunits but also to the alternative existence of a transmembrane domain or a glycolipid membrane anchor.

Acetylcholinesterase↗
PubMed1991

Lipid mediators contribute to oxygen-radical-induced airway responses in sheep.

The purpose of this study was to determine if the bronchoconstriction and airway hyperresponsiveness (AHR) resulting from aerosolized xanthine (x; 0.1%)-xanthine oxidase (xo; 4.1 U) and the subsequent production of oxygen radicals is mediated by the secondary generation of lipid mediators. In seven conscious sheep, specific lung resistance (SRL) was measured before and after x-xo challenge; approximately 30 min later when SRL had returned to baseline, airway responsiveness to carbachol was determined from dose-response curves by calculating the cumulative provocating dose of carbachol in breath units (BU, defined as one breath of a 1% wt/vol carbachol solution) that increased SRL 400% over baseline (PD400). Inhaled x-xo caused in immediate increase in SRL of 162 +/- 36% (mean +/- SE; p less than 0.05) over baseline and decreased PD400 from a baseline value of 32.5 +/- 5.0 to 16.6 +/- 1.7 BU (p less than 0.05). Pretreatment with the H2O2 scavenger, catalase (CAT,; 38 mg aerosol), methylprednisolone succinate (MS; 1 mg/kg given intravenously), the cyclooxygenase inhibitor, indomethacin (IND; 2 mg/kg given intravenously), and the PAF antagonist, WEB-2086 (3 mg/kg given intravenously) all attenuated the x-xo-induced increase in SRL (p less than 0.05); the leukotriene D4 antagonist, MK-571 (5 mg by aerosol) had no effect. All agents inhibited the x-xo-induced decrease in PD400: mean BUs were 27 after CAT, 32 after WEB-2086, 34 after IND, 31 after MS, and 25 after MK-571 (all p less than 0.05 versus x-xo alone).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Hypoxia inhibits renal thromboxane and not prostacyclin release.

The effect of hypoxia on renal prostanoid synthesis and renal function was studied. The kidneys of male Sprague-Dawley rats were cannulated and perfused in vitro with Krebs-Henseleit buffer with a pO2 of 460 (control) or 60 mm torr (hypoxia). The rate of perfusion was adjusted to maintain perfusion pressure at 100 mm Hg. The renal venous effluent was collected at 15, 30, 60, 120, 150, and 180 min and assayed for basal release of 6-keto-PGF1 alpha, PGE2, and thromboxane B2 by radioimmunoassay and inulin and sodium clearance. Prostanoid release was high at 15 min, decreased to a lower level up to 120 min, and then progressively increased after 150 min of perfusion. Hypoxia significantly decreased TxB2 release at 30, 60, 150, and at 180 min but not PGE2 or 6-keto-PGF1 alpha release. Hypoxia proportionally decreased both sodium and inulin clearances suggesting that the decrease in renal function was from decreased renal flow. The kidney responded to the altered renal function by increasing the ratio of vasodilator to vasoconstrictor prostanoids by inhibition of TxB2 release.

6-Ketoprostaglandin F1 alpha↗
PubMed1990

Addison's disease secondary to prostatic carcinoma. A case report.

Adrenal cortical insufficiency secondary to destruction of the cortex by a metastatic tumor is a rare condition. Addison's disease is usually caused by an autoimmune process or by a tuberculous infection. We report a case of adrenal insufficiency as the first clinical manifestation of a metastatic prostate carcinoma that occurred simultaneously with an active pulmonary infection by M. tuberculosis.

Addison Disease↗
PubMed1990

Cell modulation of hydrophobic tailed 16S acetylcholinesterase by intracellular calcium in rat superior cervical ganglion neurons.

In primary cell cultures of rat superior cervical ganglia (SCG) the tailed asymmetric 16S molecular form of acetylcholinesterase (AChE) possesses hydrophilic (high-salt soluble, HSS) and hydrophobic (detergent extracted, DE) variants. Hydrophobic tailed acetylcholinesterase is associated with membranes through a glycolipid anchor. In the presence of tunicamycin, an antibiotic which inhibits protein glycosylation, the cellular amount of the hydrophobic DE 16S AChE is increased. Exposure of the cells to the calcium ionophore A 23187 leads to a decrease in DE 16S AChE and a correlated increase in hydrophilic HSS 16S AChE. These results suggest the existence of an endogenous processing of tailed AChE, transforming the hydrophobic variant into an hydrophilic one controlled through glycosylation and intracellular calcium.

Acetylcholinesterase↗
PubMed1990

Induction of normal ultrastructure by CGRP treatment in dysgenic myotubes.

The calcitonin gene-related peptide (CGRP) restores an apparent normal ultrastructure in mdg/mdg muscle cells in vitro, including a normal triadic organization which is known to be essential for excitation-contraction (E-C) coupling. However, neither slow L-type Ca2+ channel activity nor E-C coupling, which are absent in mdg/mdg muscle, were re-established. These observations suggest a potential role of CGRP (and also of cAMP as the intracellular messenger) in the morphological development of the muscle fiber.

Animals↗
PubMed1990

Neuromuscular development following tetrodotoxin-induced inactivity in mouse embryos.

Developmental aspects of the neuromuscular system in mouse embryos chronically paralyzed in utero with tetrodotoxin (TTX) between embryonic days 14 and 18 were studied using biochemical and histological methods. The number of lumbar spinal motoneurons (MNs) was higher in inactive embryos than in controls suggesting a decreased motoneuron cell death. In association with the increase in MN number, choline acetyltransferase activity was significantly increased in both spinal cord and peripheral synaptic sites. Paralyzed muscles exhibited a decreased number of mature myofibers and the nuclei were centrally located. Creatine kinase activity was greatly decreased and total acetylcholine receptor and receptor cluster numbers per myofiber were significantly increased in paralyzed muscles. A similar pattern of changes occurs in the neuromuscular system of the mutant mouse muscular dysgenesis (mdg). However, in contrast to the mdg mutant, tetrodotoxin-treated muscles were similar to controls in their innervation pattern, in the ultrastructural aspects of the excitation-contraction coupling system (i.e., dyads and triads) and in the extent of dihydropyridine binding. Thus, neuromuscular inactivity is not sufficient to impair the pattern of muscle innervation or the appearance of either the triadic junctions or dihydropyridine receptors. These results indicate that alterations of dihydropyridine binding sites and triads in muscular dysgenesis cannot be accounted for by inactivity but rather must reflect a more primary defect involving the structural gene(s) regulating the development of one or more aspects of muscle differentiation.

Animals↗
PubMed1990

Hemorrhage in the upper digestive tract caused by an aorto-esophageal fistula.

The case of a 75-year-old patient admitted to our Center with hematemesis is presented. At oral endoscopy a pulsating vegetative mass was found in the distal third of the esophagus, which aroused suspicion of an aortic aneurysm fistulized to the esophagus; this was confirmed by arteriography. He underwent two surgical procedures, the aneurysm being resected and replaced by an aortic prosthesis, and esophagectomy performed. The patient died of postoperative complications. The endoscopic appearance is shown.

Aged↗